Four types of Bifidobacteria trigger autophagy response in intestinal epithelial cells
Four types of Bifidobacteria trigger autophagy response in intestinal epithelial cells
复制标题
四种双歧杆菌触发肠上皮细胞自噬反应
DOI：
10.1111/1751-2980.12179
复制
发表时间：
2014-11-01
影响因子：
3.5
通讯作者：
Hou, Xiao Hua
中科院分区：
文献类型：
--
作者：
Lin, Rong;Jiang, Ye;Hou, Xiao Hua
目的研究肠道微生物区系对肠上皮细胞(IEC)自噬激活的影响，并评价IEC对不同类型双歧杆菌的自噬反应。方法用致病性大肠杆菌(EPEC)O127：B8的脂多糖(LPS)和4种双歧杆菌的培养上清处理IEC-18细胞。用上皮伏安计测量跨皮细胞电阻(TEER)。利用串联的mCherry-GFP-LC3构建方法，通过透射电子显微镜、Lc3-II与Lc3-I的比值以及绿色荧光蛋白(GFP)和mCherry信号的持续存在来检测自噬。结果与未经处理的对照组相比，EPEC-内毒素可使IEC的TEER降低45~55%。在用双歧杆菌治疗后，在所有时间点都没有观察到这种减少。根据Lc3-II/Lc3-I的比值和透射电子显微镜，双歧杆菌可以启动IEC自噬的激活。四种双歧杆菌对自噬反应的影响无差异。与双歧杆菌相比，IEC通过自噬积累对EPEC-内毒素的反应更为强烈。经双歧杆菌和EPEC-LPS处理后，可检测到更多的mCherry(+)LC3自噬斑点和自噬基因ATG5、Atg12和Atg16的表达增加。结论双歧杆菌启动了IEC的自噬激活。Atg12-ATG5-Atg16多聚体可能参与了双歧杆菌诱导的细胞自噬的激活。
ObjectivesTo investigate the influence of gut microbiota on autophagy activation in intestinal epithelial cells (IEC) and to evaluate the IEC autophagy response to different types of Bifidobacteria.MethodsIEC-18 cells were treated with lipopolysaccharide (LPS) derived from enteropathogenic Escherichia coli (EPEC) O127:B8 and culture medium supernatants of four types of Bifidobacteria. Transepithelial electrical resistance (TEER) was measured using an epithelial voltohmmeter. Autophagy was determined by transmission electron microscopy (TEM), the ratio of LC3-II to LC3-I and the persistence of both green fluorescent protein (GFP) and mCherry signals using a tandem mCherry-GFP-LC3 construct. The expression of Atg12-Atg5-Atg16 complex was measured by quantitative real-time polymerase chain reaction.ResultsEPEC-LPS significantly diminished the TEER of IEC compared with untreated controls by 45-55%. This reduction was not observed after treated with Bifidobacteria at all time points. Bifidobacteria could initiate the activation of autophagy in IEC, based on both the ratio of LC3-II to LC3-I and TEM. There was no difference in the influence of the four types of Bifidobacteria on the autophagy response. Compared with Bifidobacteria, IEC reacted to EPEC-LPS much more intensively by autophagy accumulation. More mCherry(+)LC3 autophagic puncta and increased expressions of autophagy genes Atg5, Atg12 and Atg16 could be detected after being treated with Bifidobacteria and EPEC-LPS.ConclusionsBifidobacteria initiate autophagy activation in IEC. The Atg12-Atg5-Atg16 multimeric complex might participate in the activation of Bifidobacteria-induced cell autophagy.